HA is a non-sulphated GAG characterized by its large length and unique mode of synthesis (Fraser et al., 1997), which occurs at the plasma membrane of the cell via a hyaluronan synthase (HAS) (Weigel et al., 1997). Mammals possess three HAS genes, termed HAS1, HAS2 and HAS3, each residing at a different chromosome location (Spicer et al., 1997). Because of its mode of synthesis, HA is extruded directly into the extracellular space (Fig. 5), and is present as a coat surrounding all chondrocytes. It is likely that proteoglycan aggregate formation initially occurs in this pericellular location. The mechanism whereby the proteoglycan aggregates are released from this environment and move to the more remote parts of the extracellular matrix is not clear. In cartilage, HA size decreases and its abundance increases with age (Holmes et al., 1988). The decrease in size appears to be by degradation rather than synthesis, and could involve the action of hyaluronidases or free radicals.
While each of the three HA synthases presumably produces HA of an identical composition, there are differences in the chain length of the product and the ease with which it can be released from the cell surface (Itano et al., 1999). Thus, the properties of the extracellular matrix could differ depending on the HAS being utilized. HAS1, HAS2 and HAS3 messages are all expressed in cartilage (Recklies et al., 2001). HAS2 message has been reported to have the highest expression levels and HAS3 message the lowest levels in human chondrocytes, though the different messages exhibit variation in their regulation by growth factors and cytokines. HAS2 appears to be by far the most important synthase during embryonic development, as mice lacking expression of this gene die during mid-gestation (Camenisch et al., 2000). In contrast, those lacking the HAS1 or HAS3 genes are viable and fertile and exhibit no observable consequence on cartilage. However, it is not clear from these knockout experiments whether absence of the HAS2 gene has a direct influence on any cartilage type, as foetal development does not proceed. Recent studies on cartilage-specific HAS2 knockout mice demonstrate that the absence of HAS2 gene expression in cartilage is catastrophic; resulting in a severely abnormal skeleton and perinatal death (Fig. 6). This underlies the essential role of the hyaluronan and proteoglycan aggregates in both normal cartilage function and the chondrogenic differentiation associated with endochondral bone formation. It also illustrates that, at least in cartilage, HAS1 or HAS3 expression cannot compensate for a deficiency in HAS2.